胆固醇代谢平衡调控机制和合成课件

胆固醇代谢平衡调控

机制和合成胆固醇代谢平衡调控

机制和合成提纲胆固醇的生物学功能胆固醇相关疾病胆固醇的生物合成胆固醇在血液中的运输家族型高胆固醇血症NPC疾病胆固醇的降解与清除胆固醇代谢的负反馈调控机制—SCAP-SREBP途径胆固醇代谢的负反馈调控机制—HMGCR降解途径饮食胆固醇吸收的分子途径提纲胆固醇的生物学功能Cholesterol（胆固醇）Cholesterol（胆固醇）一、胆固醇的生物学功能MembraneFluidityBileAcidsHormones、VitaminDSynapseHedgehogModificationSignalTransduction……一、胆固醇的生物学功能MembraneFluidity哺乳动物细胞模式图哺乳动物细胞模式图膜上的微结构域膜上的微结构域Cholesterolandcholesterolderivatives

雌激素雄激素Cholesterolandcholesterolde胆固醇代谢平衡调控机制和合成课件脂肪酸、糖等提供能量（ATP）胆固醇并不提供能量脂肪酸、糖等提供能量（ATP）二、胆固醇相关疾病二、胆固醇相关疾病CholesterolisthemajorcauseofatherosclerosisCholesterolisthemajorcause心脑血管疾病恶性肿瘤肺炎、流感感染性疾病意外死亡慢性阻塞性肺病肝病及肝硬化糖尿病肾病其它43.8%22.3%FromHeJ.etal.,NEnglJMed,(2005),353:1124-1134.中国死亡原因统计心脑血管疾病恶性肿瘤肺炎、流感感染性疾病意外死亡慢性阻塞性肺蒙娜丽莎患有高胆固醇？意大利帕勒莫大学的病理解剖学教授弗兰克，研究认为蒙娜丽莎饮食不健康，患有高胆固醇症。正是眼部的脂肪瘤，造成了蒙娜丽莎这副神秘莫测的表情。画中的她（或者说达芬奇的这个模特）正在为自己体内过高的胆固醇而担忧。蒙娜丽莎患有高胆固醇？意大利帕勒莫大学的病理解胆结石老年痴呆症CholesterolandDiseasesNiemann-PickTypeCDisease糖尿病/肥胖症胆结石老年痴呆症CholesterolandDiseasFromIMSHealth,MIDAS

药品销售排行榜NameCompanyIndicationSalestoJune07($bn)LipitorPfizerHypercholesterolaemia（高胆固醇血症）13.5NexiumAstraZenecaGastroesophagealreflux（胃食管返流）,gastriculcers（胃溃疡）6.9SeretideGSKAsthma（哮喘）6.7PlavixBristol-MyersSquibbAtheroscleroticevents（动脉粥样硬化症）5.8AranespAmgenAnaemia（贫血症）5.1FromIMSHealth,MIDAS药品销售排行榜N三、胆固醇的生物合成三、胆固醇的生物合成CholesterolisnotrequiredinthedietCholesterolisanessentialmoleculebutisnotrequiredinthedietbecauseallcellscansynthesizeitfromsimpleprecursorsCholesterolisnotrequiredinCholesterolismadefromacetyl-CoAinfourstagesallofitscarbonatomsareprovidedbyasingleprecursor–acetateCholesterolismadefromacetyStage1Threeacetateunitscondensetoformasixcarbonintermediate,mevalonateStage1ThreeacetateunitsconTwomoleculesofacetate-CoAcondenseformingacetoacetyl-CoA.Acetoacetyl-CoAcondenseswithacetyl-CoAtoyieldb-hydroxy-b-methylglutaryl-CoA(HMG-CoA)Twomoleculesofacetate-CoAcThefinalstepthereductionofHMG-CoAtomevalonate,catalyzedbyHMG-CoAreductase.ThefinalstepthereductionoStage2ConversionofmevalonateintoactivatedisopreneunitsIsoprenecontainingmoleculesareimportantintermediatesincholesterolbiosynthesisStage2ConversionofmevalonatStage3Polymerizationofsix5-carbonisopreneunitstoformthe30-carbonlinearstructureofsqualene.Stage3Polymerizationofsix5Cyclizationofsqualeneformsthefourringsofthesteroidnucleus.Subsequentmodificationsleadstothefinalproduct,cholesterol.Stage4Cyclizationofsqualeneforms胆固醇代谢平衡调控机制和合成课件MostofthecholesterolmadeintheliverisexportedMuchofcholesterolsynthesistakesplaceinthe

liverMostisexportedMostofthecholesterolmadeiCholesterolisexportedinthreeforms1.Bilesalts–amphipathiccholesterolderivativesthataidlipiddigestion2.Cholesterol–tobile3.Cholesterylesters–transportedandsecretedin lipoproteinparticlestoother tissuesthatusecholesterol orarestoredintheliverCholesterolisexportedinthr四、胆固醇在血液中的运输四、胆固醇在血液中的运输CholesterylesterformationFormedintheliverConvertingcholesteroltoamorehydrophobicformCholesterylesterformationForCholesteroltransport:theproblemCholesterolandcholesterylestersareessentiallyinsolubleinwaterThesemoleculesmustbemovedfromthetissueoforigintothetissuesinwhichtheyarestoredorareconsumedCholesteroltransport:theproCholesteroltransport:thesolutionCholesterolandcholesterylestersarecarriedinthebloodplasmafromonetissuetoanotherasplasmalipoproteinsCholesteroltransport:thesolCholesterolestersentercellsby

receptormediatedendocytosisCholesterolestersentercells五、家族型高胆固醇血症LDLreceptor(LDLR)突变五、家族型高胆固醇血症LDLreceptor(LDLR)家族性高胆固醇血症(FamilialHypercholesterolemia,FH)杂合子患者血清总胆固醇较正常人高出1～2倍纯合子患者血清总胆固醇较正常人高出6～8倍杂合子患者发生率为1/500纯合子患者发生率为1/1,000,000杂合子患者男性30～40岁时，患CAD，23%患者在50岁以前死于CAD，>50%患者在60岁时明显的CAD症状；纯合子患者十几岁时，有严重的心血管事件甚至死亡家族性高胆固醇血症杂合子患者血清总胆固醇较正常人高出1～2倍LDLR突变----黄色瘤LDLR突变----黄色瘤FH患者LDLR突变----眼底脂质渗出正常人FH患者LDLR突变----眼底脂质渗出正常人六、

Niemann-PicktypeC(NPC)疾病溶酶体堆积型疾病（LysosomalStorageDisorders）胆固醇在溶酶体中堆积进行性神经细胞死亡、肝脾肿大、小脑共济失调、痴呆、语言吞咽困难、青春期之前死亡1：120，000发病，携带者1：100NPC1（大的膜蛋白）或NPC2（小的可溶蛋白）基因突变临床尝试用环化糊精进行治疗六、Niemann-PicktypeC(NPC)疾病正常细胞NPC1突变细胞正常细胞NPC1突变细胞胆固醇代谢平衡调控机制和合成课件胆固醇代谢平衡调控机制和合成课件七、胆固醇的降解与清除七、胆固醇的降解与清除DegradationofcholesterolTheringstructureofcholesterolcannotbemetabolizedtoCO2andH2OinhumansTheintactsterolringiseliminatedfromthebodyby:Conversiontobileacids,whichareexcretedinfecesSecretionofcholesterolintothebile,whichtransportsittotheintestineforeliminationDegradationofcholesterolTheSteroidhormonesareformedfromcholesterolAllsteroidhormonesarederivedformcholesterolInthecortexofadrenalglandstwoclassesofhormonesaresynthesized–mineralocorticoidsandglucocorticoidsInthemaleandfemalegonads–sexhormonesareproducedSexhormonesinclude–progesterone,androgensandestrogensSteroidhormonesareformedfr八、胆固醇代谢的负反馈调控机制（一）SCAP-SREBP途径八、胆固醇代谢的负反馈调控机制（一）SCAP-SREBP途径胆固醇代谢平衡调控机制和合成课件脂质代谢的关键蛋白质及其功能调控的临床意义LDLReceptorsLDLHMGCRSREBPPathwayHMGCRinhibitor脂质代谢的关键蛋白质及其功能调控的临床意义LDLLDLHMGInsigInsigSREBP—膜结合的转录因子WangX,BriggsMR,HuaX,YokoyamaC,GoldsteinJL,BrownMS.Nuclearproteinthatbindssterolregulatoryelementoflowdensitylipoproteinreceptorpromoter.II.Purificationandcharacterization.JBiolChem.1993Jul5;268(19):14497-504.YokoyamaC,WangX,BriggsMR,AdmonA,WuJ,HuaX,GoldsteinJL,BrownMS.SREBP-1,abasic-helix-loop-helix-leucinezipperproteinthatcontrolstranscriptionofthelowdensitylipoproteinreceptorgene.Cell.1993Oct8;75(1):187-97.WangX,SatoR,BrownMS,HuaX,GoldsteinJL.SREBP-1,amembrane-boundtranscriptionfactorreleasedbysterol-regulatedproteolysis.Cell.1994Apr8;77(1):53-62.SREBP—膜结合的转录因子WangX,BriggsM胆固醇代谢平衡调控机制和合成课件25-Hydroxycholesterol-irradiationMutantcellssurvive(25-RA)25-Hydroxycholesterol-irradiaAmphotericinB-irradiationCholesterolauxotrophs(M19)HumanfibroblasmgenomicDNAtoM19cellsHfT1M19(c)Growinlipid-deficientmediumGrowinlipid-deficientmediumHfT1M19(c)genomicDNAtoM19cellsHfT2M19(c)Growinlipid-deficientmediumHfT2M19(c)genomicDNAtoM19cellsHfT3M19(c)Inter-AluPCR,usethisprobetoscreengenomicPAClibrary,transfectPACcloneintoM19,sequencePACcloneandcomparewithEST.AmphotericinB-irradiationChS2PcDNAtoCHOcellsBrieflyincubatedwithLDL,AmphotericinBselection-irradiationLowfluorescentLDLuptake-irradiationSRD-12BS2PcDNAtoCHOcellsBrieflyiDeBose-BoydRA,BrownMS,LiWP,NohturfftA,GoldsteinJL,EspenshadePJ.Transport-dependentproteolysisofSREBP:relocationofsite-1proteasefromGolgitoERobviatestheneedforSREBPtransporttoGolgi.Cell.1999Dec23;99(7):703-12.NohturfftA,YabeD,GoldsteinJL,BrownMS,EspenshadePJ.RegulatedstepincholesterolfeedbacklocalizedtobuddingofSCAPfromERmembranes.Cell.2000Aug4;102(3):315-23.SterolRegulatedTransportofSCAPDeBose-BoydRA,BrownMS,LiWPurificationSchemeforSCAP-interactingProteinsYangetal.,Cell(2002)489-500PurificationSchemeforSCAP-iTwoInsigs:Insig-1andInsig-2A.SequenceAlignmentB.HydropathyPlotYabeetal.,PNAS(2002)12753-8TwoInsigs:Insig-1andInsig-InsigInsig九、胆固醇代谢的负反馈调控机制（二）HMGCR降解途径九、胆固醇代谢的负反馈调控机制（二）HMGCR降解途径FromHeJ.etal.,NEnglJMed,(2005),353:1124-1134.andIMSHealth,MIDAS

药品销售排行榜NameCompanyIndicationSalestoJune07($bn)LipitorPfizerHypercholesterolaemia（高胆固醇血症）13.5NexiumAstraZenecaGastroesophagealreflux（胃食管返流）,gastriculcers（胃溃疡）6.9SeretideGSKAsthma（哮喘）6.7PlavixBristol-MyersSquibbAtheroscleroticevents（动脉粥样硬化症）5.8AranespAmgenAnaemia（贫血症）5.1FromHeJ.etal.,NEnglJMeHMG-CoAReductase(HMGCR):theRate-LimitingEnzymeinCholesterolBiosyntheticPathway(HMGCR)HMG-CoAReductase(HMGCR):theProposedModelforINSIG-mediatedRegulationof

HMGCoAReductaseandSCAPProposedModelforINSIG-mediaSterol-RegulatedDegradationofHMG-CoAReductaseisBlockedbyInhibitorsofthe26SProteasome

Sterol-RegulatedDegradationoWorkingHypothesisforSterol-RegulatedDegradationofHMG-CoAReductaseWorkingHypothesisforSterol-Sterol-StimulatedUbiquitinaionofHMGCRRequiresInsigSever,Song,Yabe,etal.,JBC,2003Sterol-StimulatedUbiquitinaioAminoAcidSequenceoftheHMG-CoAReductase

MembraneDomainAminoAcidSequenceoftheHMGLysines89and248areRequiredfortheSterol-RegulatedUbiquitinationandDegradationofHMGCR

Lysines89and248areRequireStrategyforPurifyingtheE3ofHMG-CoAReductaseStrategyforPurifyingtheE3E3ActivityCo-immunoprecipitateswithInsig-1inSterol-TreatedCellsE3ActivityCo-immunoprecipitaIdentificationofgp78andVCPasInsig-1-associatingproteinsIdentificationofgp78andVCP胆固醇代谢平衡调控机制和合成课件MolecularPathwayforSterol-regulatedDegradationofHMGCRSongetal.,MolCell,2005Whatareotherproteinsinvolvedinthispathway?MolecularPathwayforSterol-rIdentificationofUfd1asagp78interactingproteinIP:Endogenousgp78IP:Endogenousgp78Transfect&CoIPIP:Flag-Ufd1ABCIdentificationofUfd1asagpUfd1isrequiredfortheubiquitinationofendogenousHMGCRUfd1isrequiredfortheubiquUfd1enhancestheE3activityofgp78invitroUfd1enhancestheE3activityDualrolesofUfd1:EnhancingE3activityandPromotingdegradationCaoetal.,CellMetab,2007DualrolesofUfd1:Enhancing十、饮食胆固醇吸收的分子途径十、饮食胆固醇吸收的分子途径(HMGCR)DenovoCholesterolSynthesisisaEnergy-ConsumingProcess18

Acetyl-CoACholesterol27

NADPH11

O218

ATP(HMGCR)DenovoCholesterolSynCholesterolBiosynthesisandAbsorptioninHuman300-500mgDietaryAbsorptionBiosynthesis600-900mg~1000mgBiliaryre-absorptionCholesterolBiosynthesisandADietaryCholesterolAbsorptionbyIntestinalEnterocytesCholesterolEsterHydrolysisCholesterolCholesterolCholesterolEsterChylomicronIntestinallumenLymphACAT2EREnterocyteTightJunctionEzetimibe2001CholesterolabsorptioninhibitorEzetimibeNPC1L12004IdentificationofNPC1L1DietaryCholesterolAbsorptionHumanNiemannPickC1

Like1(NPC1L1)HumanNiemannPickC1Like1(NPC1L1mediatesthere-absorptionofcholesterolfrombileApicalHepatocytesBasolateralApicalNPC1L1mediatesthere-absorptNPC1L1recyclesbetweenPMandERCrespondingtocholesterollevelABNPC1L1recyclesbetweenPMandOverexpressionofNPC1L1increasesfreecholesteroluptakeOverexpressionofNPC1L1increKnockingdownofNPC1L1proteininL02cellsattenuatestheuptakeoffreecholesterolKnockingdownofNPC1L1proteiStructuresofdifferentsterolsSterolAbsorption:<5%plantsterols>50%cholesterolStructuresofdifferentsterolSterol-specificityforNPC1L1-mediatedinternalizationABSterol-specificityforNPC1L1-Hypothesis:NPC1L1mediatescholesterolupta